GSM7049490: S53-P002-C-240m; Homo sapiens; RNA-Seq
Total RNA was isolated from PBMC samples using the RNeasy Plus Mini Kit (QIAGEN GmbH, Hilden, Germany) and eluted in 60µL RNase-Free water 1μg total RNA was used for library preparation. Poly(A) mRNA isolation was performed using Poly(A) mRNA Magnetic Isolation Module. The mRNA fragmentation and priming were performed using First Strand Synthesis Reaction Buffer and Random Primers. First strand cDNA was synthesized using ProtoScript II Reverse Transcriptase and the second-strand cDNA was synthesized using Second Strand Synthesis Enzyme Mix. The purified double-stranded cDNA by beads was then treated with End Prep Enzyme Mix to repair both ends and add a dA-tailing in one reaction, followed by a T-A ligation to add adaptors to both ends. Size selection of Adaptor-ligated DNA was then performed using beads, before PCR amplification and validation of PCR products by Qsep100 (Bioptic, Taiwan, China) and quantification by Qubit3.0 Fluorometer (Invitrogen, Carlsbad, CA, USA)